ABSTRACT (Functional Genomics Core) Antimicrobial resistance (AMR) and its impact have been recognized by the WHO, CDC, FDA, and NIH as one of the most important public health threats facing society today. Vancomycin-resistant enterococci (VRE), extended spectrum ?-lactamase/carbapenemase-producing Enterobacteriaceae (ESBL-E/CRE), and Clostridiodes difficile are of particular interest as they disproportionately affect immunocompromised and severely ill patients. The CDC has designated VRE as a serious threat and both ESBL-E/CRE and C. difficile as urgent threats. Each of these pathogens is able to colonize and infect the gut, with disruption of the protective gut microbiome by antibiotics a leading risk factor for infection. The overarching hypothesis of the DYNamics of colonizAtion and infection by Multidrug-resIstant paThogens in immunocompromisEd patients program (DYNAMITE) is that patient susceptibility to nosocomial acquisition, gut colonization, and subsequent infection by pathogens is critically dependent on functional microbiota-pathogen interactions that determine disease progression and clinical outcomes. Importantly, we posit that shotgun metagenomics data coupled with inferred metabolic potential is not enough to predict and interpret susceptibility to colonization and infection in a dysbiotic patient, and a combination of genomics, metagenomics, metabolomics, and metaproteomics is necessary to elucidate the complex interplay between the pathogen, the microbiome, and the host. To enable this multi-omic approach, the Functional Genomics Core (FGC) will provide a central resource to all three projects contained within this application, providing facilities and expertise for whole genome sequencing, metagenomics, metaproteomics, and metabolomics. Additionally, leveraging the robust bioinformatics infrastructure housed within the FGC, we will provide the application of existing analytical pipelines and disease classifiers?as well as the development of novel pipelines?that will integrate and facilitate the comprehensive analyses of the data generated for each individual project. As an example, we have built a high-throughput pipeline for the generation and annotation of reference-quality fully circularized bacterial chromosomes and accompanying plasmids, and with accompanying metaproteomics data have mapped 92% of protein signatures back to the source gene. Overall, to achieve the scientific goals outlined in this P01 application, the FGC will undertake three aims: i) coordinate the transfer of pure microbiological isolates and patient stool samples from the study sites to the FGC; ii) complete genomic, metagenomic, metaproteomic, and metabolomic processes of each project; and iii) assist in the analysis and integration of the multi-omic data generated under each project. The FGC provides a unique and unparalleled ability to combine multiple omics data types to create a truly multi-omic assessment of the complex interplay between VRE, ESBL-E/CRE, and C. difficile, the gut microbiome, and the host metabolome, facilitating the goals of this DYNAMITE program.